ASTM E1303-1995(2005) Practice for Refractive Index Detectors Used in Liquid Chromatography《液相色谱法用折射指数检测器规程》.pdf

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1、Designation: E 1303 95 (Reapproved 2005)Standard Practice forRefractive Index Detectors Used in Liquid Chromatography1This standard is issued under the fixed designation E 1303; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revision, the y

2、ear of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon (e) indicates an editorial change since the last revision or reapproval.1. Scope1.1 This practice covers tests used to evaluate the perfor-mance and to list certain descriptive specifications of

3、 arefractive index (RI) detector used as the detection componentof a liquid chromatographic (LC) system.1.2 This practice is intended to describe the performance ofthe detector both independent of the chromatographic system(static conditions, without flowing solvent) and with flowingsolvent (dynamic

4、 conditions).1.3 The values stated in SI units are to be regarded as thestandard.1.4 This standard does not purport to address all of thesafety concerns, if any, associated with its use. It is theresponsibility of the user of this standard to establish appro-priate safety and health practices and de

5、termine the applica-bility of regulatory limitations prior to use.2. Referenced Documents2.1 ASTM Standards:2E 386 Practice for Data Presentation Relating to High-Resolution Nuclear Magnetic Resonance (NMR) Spectros-copy3. Significance and Use3.1 Although it is possible to observe and measure each o

6、fseveral characteristics of a detector under different and uniqueconditions, it is the intent of this practice that a complete set ofdetector test results should be obtained under the same oper-ating conditions. It should also be noted that to specifycompletely a detectors capability, its performanc

7、e should bemeasured at several sets of conditions within the useful rangeof the detector.3.2 The objective of this practice is to test the detector underspecified conditions and in a configuration without an LCcolumn. This is a separation independent test. In certaincircumstances it might also be ne

8、cessary to test the detector inthe separation mode with an LC column in the system, and theappropriate concerns are also mentioned. The terms and testsdescribed in this practice are sufficiently general so that theymay be adapted for use at whatever conditions may be chosenfor other reasons.4. Noise

9、, Drift, and Flow Sensitivity4.1 Descriptions of Terms Specific to This Standard:4.1.1 short term noisethis noise is the mean amplitude inrefractive index units (RIU) for random variations of thedetector signal having a frequency of one or more cycles perminute. Short term noise limits the smallest

10、signal detectableby an RI detector, limits the precision attainable, and sets thelower limit on the dynamic range. This noise corresponds toobserved noise of the RI detector only. (The actual noise of theLC system may be larger or smaller than the observed value,depending upon the method of data col

11、lection, or signalmonitoring of the detector, since observed noise is a function ofthe frequency, speed of response and the band width of therecorder or other electronic circuit measuring the detectorsignal.)4.1.2 long term noisethis noise is the maximum ampli-tude in RIU for random variations of th

12、e detector signal withfrequencies between 6 and 60 cycles per h (0.1 and 1.0 cyclesper min). It represents noise that may be mistaken for alate-eluting peak. This noise corresponds to the observed noiseonly and may not always be present.4.1.3 driftthe average slope of the long term noise enve-lope e

13、xpressed in RIU per hour as measured over a period of1h.4.1.4 staticrefers to the noise and drift measured underconditions of no flow.4.1.5 dynamicrefers to the noise and drift measured at aflow rate of 1.0 mL/min.4.1.6 flow sensitivitythe rate of change of signal displace-ment (in RIU) vs flow rate

14、 (in mL/min) resulting from stepchanges in flow rate calculated at 1 mL/min as described in4.3.12.4.2 Test Conditions:4.2.1 The same test solvent must be used in both sample andreference cells. The test solvent used and its purity should bespecified. Water equilibrated with the laboratory atmosphere

15、1This practice is under the jurisdiction of ASTM Committee E13 on MolecularSpectroscopy and Chromatography and is the direct responsibility of SubcommitteeE13.19 on Chromatography.Current edition approved Feb. 1, 2005. Published March 2005. Originallyapproved in 1989. Last previous edition approved

16、in 2000 as E 1303 95 (2000).2For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM Customer Service at serviceastm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.1Copyright ASTM International, 10

17、0 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.containing minimum impurities is the preferred test solvent formeasuring noise and drift. Water for this purpose (preferablypurified by distillation, deionization, or reverse osmosis)should be drawn, filtered through a

18、 0.45-m filter, and allowedto stand in a loosely covered container for several hours atambient temperature in the laboratory in which testing is to becarried out. This will ensure complete equilibration of thewater with the gases in the laboratory atmosphere.NOTE 1It is essentially impossible to mai

19、ntain a constant RI value ofde-gassed water and of very dilute samples in de-gassed water. This is dueto the fact that the difference in refractive index between completelyde-gassed water and atmosphere-equilibrated water is 1.5 3 106RIU.3Thus, small differences in the concentration of dissolved gas

20、es betweenthe sample and the trapped reference can lead to significant errors inmeasurement of solutions where the expected difference in RI due tosolute is of the order of 106RIU or less. Therefore, in order to minimizeerror in determining samples with small RIU differences between them,atmosphere-

21、equilibrated water (5.2.1) is recommended as the solvent fordetermining linearity and minimum detectability (Section 5).4.2.2 The detector should be located at the test site andswitched on at least 24 h prior to the start of testing. Somedetectors provide an oven to thermostat the optics assembly.Th

22、e oven should be set at a suitable temperature, following themanufacturers recommendations, and this temperature shouldbe noted and maintained throughout the test procedures.4.2.3 Linearity and speed of response of the recorder orother data acquisition device used should be such that it doesnot dist

23、ort or otherwise interfere with the performance of thedetector.4If additional amplifiers are used between the detectorand the final readout device, their characteristics should alsofirst be established.4.3 Methods of Measurement:4.3.1 Connecta1m(39.37 in.) length of clean, dry,stainless steel tubing

24、 of 0.25 mm (0.009 to 0.01 in.) insidediameter in place of the analytical column. The tubing can bestraight or coiled to minimize the space requirement. Thetubing should terminate in standard low dead volume fittings toconnect with the detector and to the pump. Commercialchromatographs may already c

25、ontain some capillary tubing toconnect the pump to the injection device. If this is of a similardiameter to that specified, it should be included in the 1.0 mlength; if significantly wider, it should be replaced for this test.4.3.2 Repeatedly rinse the reservoir and chromatographicsystem, including

26、the detector, with the test solvent prepared asdescribed in 4.2.1, until all previous solvent is removed fromthe system. Fill the reservoir with the test solvent.4.3.3 Thoroughly flush the reference cell with the samesolvent; keep the reference cell static.4.3.3.1 It may be necessary to flush both s

27、ample andreference cells with an intermediate solvent (such as methanolor acetone), if the solvent previously used in the system isimmiscible with the test solvent.4.3.4 Allow the chromatographic system to stabilize for atleast 60 min without flow. The detector range, Note 2, shouldbe set such that

28、the amplitude of short term noise may be easilymeasured. Ideally, the output should contain no filtering of thesignal. If the filtering cannot be turned off, the minimum timeconstant should be set and noted in the evaluation. Manuals ormanufacturers should be consulted to determine if time con-stant

29、 and detector range controls are coupled, and informationshould be obtained to determine if they can be decoupled fortesting. Set the recorder zero to near mid-scale. Record at least1 h of baseline under these static conditions, during which timethe ambient temperature should not change by more than

30、 2C.NOTE 2RI detectors will have one or more controls labeled attenua-tion, range, sensitivity, and scale factor. All are used to set the full scalerange (in RIU) of an output display device such as a strip chart recorder.4.3.5 Draw pairs of parallel lines, each between12 to 1 minin length, to form

31、an envelope of all observed randomvariations over any 15-min period (Fig. 1). Draw the parallellines in such a way as to minimize the distance between them.Measure the distance perpendicular to the time axis betweenthe parallel lines. Convert this value to RIU (5.2.9). Calculatethe mean value over a

32、ll the segments; this value is the staticshort term noise.4.3.6 Now mark the center (center of gravity) of eachsegment over the 15-min period of the short term noise3Munk, M. N., Liquid Chromatography Detectors, (T. M. Vickrey, Ed.), MarcelDekker, New York and Basel, 1983, pp. 165204.4Bonsall, R. B.

33、, “The Chromatography SlaveThe Recorder,” Journal of GasChromatography, Vol 2, 1964, pp. 277284.FIG. 1 Examples for the Measurement of Short Term Noise, LongTerm Noise and DriftE 1303 95 (2005)2measurement. Draw a series of parallel lines to these centers,each 10 min in length (Fig. 1), and choose t

34、hat pair of lineswhose distance apart perpendicular to the time axis is greatest.This distance is the static long term noise.4.3.7 Draw the pair of parallel lines, over the1hofmeasurement, that minimizes the distance perpendicular to thetime axis between the parallel lines. The slope of either line,

35、measured in RIU/h, is the static drift.4.3.8 Set the solvent delivery system to a flow rate that haspreviously been shown to deliver 1.0 mL/min under the sameconditions of capillary tubing, solvent, and temperature. Allowat least 15 min to stabilize. Set the recorder zero nearmid-scale. Record at le

36、ast1hofbaseline under these flowingconditions, during which time the ambient temperature shouldnot change by more than 2C.4.3.9 Draw pairs of parallel lines, measure the perpendiculardistances, and calculate the dynamic short term noise, in themanner described in 4.3.5 for the static short term nois

37、e.4.3.10 Make the measurement for the dynamic long termnoise following the procedure outlined in 4.3.6.4.3.11 Draw the pair of parallel lines in accordance with4.3.7. The slope of this line is the dynamic drift.4.3.12 Stop the chromatographic flow.Allow at least 15 minfor re-equilibration. Set the r

38、ecorder at about 5 % of full scaleand leave the detector range setting at the value used for thenoise measurements. Set the solvent delivery system at a flowrate of 0.5 mL/min. Run for 15 min, or more if necessary forre-equilibration, at a slow recorder speed. Increase the flow rateto 1.0 mL/min and

39、 record for 15 min or more. Run at 2.0, 4.0,and 8.0 mL/min if the pressure flow limit of the chromato-graphic system is not exceeded. If necessary, adjust the detectorrange to maintain an on-scale response.4.3.13 Draw a horizontal line through the plateau producedat each flow rate, after a steady st

40、ate is reached (Fig. 2).Measure the vertical displacement between these lines, andexpress in RIU (5.2.9). Plot these values versus flow rate. Drawa smooth curve connecting the points and draw a tangent at 1mL/min (Fig. 3). Express the slope of the line as the flowsensitivity in RIU min/mL. It is pre

41、ferred to give the numericalvalue and show the plot as well.5. Minimum Detectability, Linear Range, DynamicRange, and Calibration5.1 Descriptions of Terms Specific to this Standard:5.1.1 minimum detectabilitythat concentration of a spe-cific solute in a specific solvent that gives a signal equal tot

42、wice the static short-term noise.5.1.1.1 DiscussionThe static short-term noise is a mea-surement of peak-to-peak noise. A statistical approach to noisesuggests that a value of three times the rms (root-mean-square)noise would ensure that any value outside this range would notbe noise with a confiden

43、ce level of greater than 99 %. Sincepeak-to-peak noise is approximately five times the rmsnoise,4,5the minimum detectability defined in this practice is amore conservative estimate. Minimum detectability, as definedin this practice, should not be confused with the limit ofdetection in an analytical

44、method using a refractive indexdetector.5.1.2 sensitivity (response factor)the signal output perunit concentration of the test substance in the test solvent, inaccordance with the following relationship:S 5 R/C (1)where:S = sensitivity (response factor), RIUL/g,R = measured detector response, RIU, a

45、ndC = concentration of the test substance in the test solventg/L.5.1.3 linear rangethe range of concentrations of the testsubstance in the test solvent, over which the sensitivity of thedetector is constant to with 5 % as determined from the5Blair, E. J., Introduction to Chemical Instrumentation, Mc

46、Graw-Hill, NewYork, NY, 1962, and Practice E 386.FIG. 2 Example for the Measurement of Flow SensitivityFIG. 3 Example of Plot for Calculation of Flow SensitivityE 1303 95 (2005)3linearity plot specified in 5.2.13. The linear range may beexpressed in three different ways:5.1.3.1 As the ratio of the u

47、pper limit of linearity obtainedfrom the linearity plot, and the minimum linear concentration,both measured for the same test substance in the same testsolvent as follows:L.R. 5 Cmax/Cmin(2)where:L.R. = linear range of the detector,Cmax= upper limit of linearity obtained from the linearityplot, g/L,

48、 andCmin= minimum linear concentration, g/L, as defined in5.2.13.1, the minimum linear concentration shouldalso be stated.5.1.3.2 By giving the minimum linear concentration and theupper limit of linearity (for example, from 8.72 3 103g/L to8.72 3 101g/L).5.1.3.3 By giving the linearity plot itself,

49、with the minimumlinear concentration and the upper limit of linearity indicatedon the plot.5.1.4 Dynamic RangeThat range of concentrations of thetest substance in the test solvent, over which an incrementalchange in concentration produces an incremental change indetector signal. The upper limit is the highest concentration atwhich a slight further increase in concentration will give anobservable increase in detector signal. The dynamic range isthe ratio of these upper and lower limits. The dynamic range islarger than or equal to the linear range, but obvi